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INNOVATIVE METHODOLOGY

Conditional gene expression in renal collecting duct epithelial cells: use of the inducible Cre-lox system

¹Institut National de la Santé et de la Recherche Médicale U478, Federative Institute of Research 02, Bichat Medical School, 75870 Paris Cedex 18; and ²Centre National de la Recherche Scientifique UMR 144, Institut Curie, 75006 Paris, France

Submitted 20 August 2002 ; accepted in final form 27 July 2003

ABSTRACT

The renal collecting duct plays a key role in control of ion and fluid homeostasis. Genes encoding for ion transporters, hormone receptors, or regulatory proteins specifically expressed in the collecting duct are mutated in several genetic diseases with altered blood pressure. Suitable cellular models expressing genes in a conditional way should represent attractive systems for structure-function analyses and generation of appropriate physiopathological models of related diseases. However, generation of such systems remains laborious and quite inefficient. We adapted and improved a conditional Cre-lox-inducible system in the highly differentiated aldosterone-sensitive rat cortical collecting duct (RCCD2) cell line. The inducible MerCreMer recombinase allowed tight control and high levels of transgene expression, whereas flanking a selection marker with two loxP sites strongly improved the selection procedure. We have used this system to conditionally express an enhanced green fluorescent protein-tagged human mineralocorticoid receptor. In the future, this will allow structure-function analyses as well as mineralocorticoid receptor trafficking studies in these epithelial cells, which retain the features of the native collecting duct. Improvements in the conditional Cre-lox expression system have potentially wide applications in other epithelial or nonepithelial cell lines.

Cre recombinase; rat cortical collecting duct 2 cells; mineralocorticoid receptor; aldosterone

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